This invention relates primarily to the use of miniaturization techniques to detect and monitor and control changes on a very small scale. More particularly, it relates to the use of micromachining techniques and micromachinery to provide highly sensitive detectors for use in measuring and selectively controlling changes occurring on a molecular level.
One of the most essential requirements for research and diagnostic protocols is the ability to detect and monitor chemical, physical, and biological reactions. There is also a need for sensors and sensor assemblies capable of detecting substances that may be present in very low concentrations. In many operations, gross observation and assays, such as titration procedures, suffice to determine whether a reaction has occurred and the extent to which it has proceeded. Especially where the quantities of reactants are large, the type of reaction is well-known, or a reaction causes a change in state or other characteristic, such as forming a precipitant, gross observation may be sufficient to monitor, characterize, and control a reaction.
In other cases, however, such observation may not be possible, or may not convey the necessary information. For certain types of basic research and diagnostic testing, and in many sensor platform applications, current monitoring or control methods and apparatus are not sensitive enough and may not be capable of detecting the reaction. Moreover, different types of information may be sought that cannot be obtained through testing of an end-product. At the same time it may be, for example, that one or more of the reactants is present in only a very small quantity, or that the end product has not been sufficiently characterized for a suitable assay to have been created.
Also, a particular project may require simultaneous monitoring of many reactions. More difficult still, the project may require such monitoring of many different types of reactions. Biocatalysts such as enzymatic proteins catalyze chemical reactions and provide specificity. Another example is the testing of pharmaceutical chemicals. It can be of great utility to sense when a particular formulation causes even the beginning of a desired reaction, or which of two or more closely related formulations drives a reaction further in the desired direction. Monitoring such activity can identify promising structures in the development of new pharmaceuticals.
The types of reactions mentioned above are only examples of the wide variety of types of reactions it is desirable to monitor and control. These types of reactions, however, are exceedingly difficult to detect and analyze with current techniques. The variety in the possible interactions, the variety in the degree to which the interactions progress, and the vast differences in the molecules to be surveyed prevent simple schemes. Signal transduction techniques such as single tagging schemes using fluorescent, radioactive, or electrochemical labels are not possible or useful. Other techniques with wide applicability and adaptability to parallel analysis requirements are few and undeveloped.
One analytic technique being used currently involves calorimetric sensing. With the available methodologies, however, calorimetric measurements require expensive analytical-grade instruments. These are not suitable or practicable for use in routine diagnostics and for other needs, including certain types of research.
One approach in calorimetric measuring has been to use packed biomolecule reactor beds. These are created by immobilizing the molecule of interest, such as an enzyme or antibody, on a structural support which is in turn placed in a chromatography column. This achieves a high process reactant-to-volume ratio to facilitate temperature measurements. A flowing stream containing the analyte is passed through the column, while thermal sensors record the inflow and outflow temperatures. This technique can only work, however, where there are available large amounts of both the reactant and the analyte.
Another early technique developed a so-called enzyme thermistor, formed by integrating an enzyme and a thermistor-type thermal probe. This technique was workable for enzymatically catalyzed reactions producing large amounts of heat. The thermistor has poor sensitivity and required protective sheathing to prevent the heat from being rapidly dissipated to the in surroundings. Thus the response time of the probe was unsatisfactory, the probe was difficult to construct, and the types of reactions for which it was appropriate were few.
Almost all sensors and platforms for calorimetric sensing are relatively large, and require large amounts of reactants and/or analytes. They are also limited to relatively high energy reactions, which are capable of generating enough heat to be detected and/or measured.
More recently, some miniaturization has been accomplished. Xie et al., Sensors and Actuators, B6: 127 (1992) have proposed immobilizing an enzyme onto a micromachined channel. The channel itself is constructed on a planar silicon surface. Measurements, however, were made using conventional thermal sensors placed outside the device.
Bataillard et al. (Biosensors and Bioelectronics 8, 89-98 (1993)) and Towe and Guilbeau (Biosensors and Bioelectronics 11(3), 247-252 (1996)) have integrated enzymes with micromachined thermopiles. While thermopiles are capable of sensing very small changes in temperature, they are not capable of sensing reactions when only a very small number or small concentration of reactants is involved. Joseph et al. (Electronic Design, 121-134 (1997)) has proposed using micromachined structures with thin platinum films for use in calorimetric gas sensing, and Berger et al., in related co-pending application Ser. No. 09/039,707, filed Mar. 16, 1998 and commonly assigned, have used a micromachined bimetallic cantilever for sensing physical transitions. These sensors can be very useful, but are subject and responsive to mechanical forces such as flow that are not necessarily indicative of temperature.
While useful for certain applications, these techniques do not provide the sensitivity, response times, and other characteristics enabling use thereof in most research and diagnoses. There is thus room for improvement in the art.
It is an object of this invention to provide a micromachined apparatus capable of detecting and measuring very small calorimetric changes.
It is also an object of this invention to provide a method for sensing enthalpic heats of reactions on a very small scale.
It is moreover an object of this invention to provide an array of micromachined thermal sensors, the array capable of sensing different types of enthalpic reactions for different types of molecules.
It is another object of this invention to provide a method and apparatus for detecting and measuring small changes to biomolecules caused by interaction thereof with other molecules or with environmental conditions.
An additional object of this invention is to provide a relatively inexpensive sensor, singly or in arrays, capable of sensing reactions in the presence of minute amounts of reactants and/or analytes.
Another object of the invention is to provide a microminiaturized, thermally isolated sensor or an array of microminiaturized thermally isolated sensors capable of detecting enthalpic changes resulting from reactions occurring in a wide variety of environments.
It is likewise an object of the invention to provide a sensor or array that enables detection of the presence of one or more particular analytes in an environment containing a relatively large number of differing analytes.
These and other objects of the invention are achieved by providing an apparatus for measuring enthalpic changes on a very small scale consisting of a thermally isolated mass; detector means located on said thermally isolated mass for sensing temperature changes adjacent said thermally isolated mass; a molecular layer adjacent said detector means, said molecular layer comprising at least one reactant of an enthalpic reaction; and an output means operatively connected to said detector means for providing an indication of said temperature changes.
These and other objects of the invention are also achieved by providing a method for detecting enthalpic changes on a very small scale consisting of micro-machining a substrate to form a thermally isolated mass; embedding in said thermally isolated mass at least one detector means responsive to changes in enthalpy and capable of providing an output indicative of enthalpic changes; coating said detector means with a molecular layer wherein the molecules of said layer comprise at least one reactant of an enthalpic reaction; and monitoring said output to detect and measure enthalpic changes.